Tumor Blood Research Articles

Impacts of Combining Pulsed Low Dose Rate Radiotherapy (PLDR) and Anti-PD-1 Antibody on Tumor Growth and Its Mechanism for Lung Cancer in Mice

As an emerging anti-tumor strategy, Immunotherapies has been approved for the treatment of a variety of tumors. Recent data suggest that the efficacy of radiotherapy in various cancers can be augmented when combined with immune checkpoint blockade. Then, pulsed low dose rate radiotherapy (PLDR) is a new radiotherapy segmentation method. Therefore, this study investigated the inhibitory effect of PLDR combined with anti-PD-1 antibody on lung cancer in mice and its impact on tumor immune microenvironment. By transplanting murine LLC cells into the right leg of C57BL/6 mice with immune activity, a transplanted subcutaneous tumor model was established. The mice were randomly divided into five groups: control, conventional radiotherapy (RT)±anti-PD-1 antibody (mAb),PLDR±anti-PD-1 antibody (mAb). RT was delivered as a dose of 5 × 2 Gy whereas PLDR involved delivering a dose of 2 Gy as 10 pulses of 0.2 Gy, each 3 minutes apart and lasting for 5 days. Anti-PD-1 antibody and isotype control were administered intraperitoneally once every three days at a dose of 5mg/kg, three times in total. The tumor, blood and tumor-draining lymph nodes (TDLNs) were harvested after treatment, and a single cell suspension was prepared for flow cytometry to analyze the changes in the immune microenvironment of the tumor tissue, the expression of PD-L1. PD-1 and the activated systemic immune response; Finally, this study explored a mechanism able to explain the observed synergy of combined therapy. PLDR combined with anti-PD-1 antibody can better inhibit the growth of tumor than RT,PLDR and RT combined group. Survival analysis demonstrated a statistically significant advantage for PLDR+anti-PD-1 than other groups. Median survival with PLDR+anti-PD-1mAb was 63d compared with 54d with RT+anti-PD-1mAb,41d with RT and 40d with PLDR and 33 with control. Meanwhile both RT and PLDR induced up-regulation of PD-L1 expression on tumor surface and PD-1 expression on lymphocytes. Then, the frequency of CD4+,CD8+T cells were higher in the PLDR combined treatment group in tumor blood and draining lymph nodes, and synergistically reduce the local accumulation of tumor-infiltrating myeloid-derived suppressor cells (MDSCs) than others. Finally, the main reason for the better tumor inhibition effect of PLDR combined group is that it upgrades the number and activity of CD8+T cells in tumor. PLDR combined with anti-PD-1 antibody can result in better tumor growth inhibition and significantly delay the survival time of mice, which was mainly through the cytotoxic T cell-dependent mechanism, meanwhile increasing the infiltration of CD4 + and CD8 +T-cells in tissues.

Induced Vascular Normalization-Can One Force Tumors to Surrender to a Better Microenvironment?

Immunotherapy has changed the way many cancers are being treated. Researchers in the field of immunotherapy and tumor immunology are investigating similar questions: How can the positive benefits achieved with immunotherapies be enhanced? Can this be achieved through combinations with other agents and if so, which ones? In our view, there is an urgent need to improve immunotherapy to make further gains in the overall survival for those patients that should benefit from immunotherapy. While numerous different approaches are being considered, our team believes that drug delivery methods along with appropriately selected small-molecule drugs and drug candidates could help reach the goal of doubling the overall survival rate that is seen in some patients that are given immunotherapeutics. This review article is prepared to address how immunotherapies should be combined with a second treatment using an approach that could realize therapeutic gains 10 years from now. For context, an overview of immunotherapy and cancer angiogenesis is provided. The major targets in angiogenesis that have modulatory effects on the tumor microenvironment and immune cells are highlighted. A combination approach that, for us, has the greatest potential for success involves treatments that will normalize the tumor's blood vessel structure and alter the immune microenvironment to support the action of immunotherapeutics. So, this is reviewed as well. Our focus is to provide an insight into some strategies that will engender vascular normalization that may be better than previously described approaches. The potential for drug delivery systems to promote tumor blood vessel normalization is considered.

Thermal analysis of different shape nanoparticles on hyperthermia therapy on breast cancer in a porous medium: A fractional model

Cancer is clearly a major cause of disease and fatality around the world, yet little is known about how it starts and spreads. In this study, a model in mathematical form of breast cancer guided by a system of (ODE’S) ordinary differential equations is studied in depth to examine the thermal effects of various shape nanoparticles on breast cancer hyperthermia therapy in the existence of a porous media with fractional derivative connection, when utilizing microwave radiative heating. The unsteady state is determined precisely using the Laplace transform approach to crop a more decisive examination of temperature dissemination of blood temperature inside the breast tissues. Durbin's and Zakian's techniques are used to find Laplace inversion. Mild temperature hyperthermia is used in the treatment, which promotes cell death by increasing cell nervousness to radiation therapy and flow of blood in tumor. In the graphical findings, we can witness the distinct behavior of hyperthermia therapy on tumor cells by applying various metabolic heat generation rates across various time intervals to attain the optimal therapeutic temperature point. Particularly, we used graphs to visualize the behavior of different Nanoparticles with different shaped during hypothermia therapy. In comparison to other nanoparticles and shapes, it demonstrates that gold nanoparticles with a platelet shape are the best option for improving heat transmission. Which assess of heat transfer up to 16.412%.

High prevalence of somatic PIK3CA and TP53 pathogenic variants in the normal mammary gland tissue of sporadic breast cancer patients revealed by duplex sequencing

The mammary gland undergoes hormonally stimulated cycles of proliferation, lactation, and involution. We hypothesized that these factors increase the mutational burden in glandular tissue and may explain high cancer incidence rate in the general population, and recurrent disease. Hence, we investigated the DNA sequence variants in the normal mammary gland, tumor, and peripheral blood from 52 reportedly sporadic breast cancer patients. Targeted resequencing of 542 cancer-associated genes revealed subclonal somatic pathogenic variants of: PIK3CA, TP53, AKT1, MAP3K1, CDH1, RB1, NCOR1, MED12, CBFB, TBX3, and TSHR in the normal mammary gland at considerable allelic frequencies (9 × 10−2– 5.2 × 10−1), indicating clonal expansion. Further evaluation of the frequently damaged PIK3CA and TP53 genes by ultra-sensitive duplex sequencing demonstrated a diversified picture of multiple low-level subclonal (in 10−2–10−4 alleles) hotspot pathogenic variants. Our results raise a question about the oncogenic potential in non-tumorous mammary gland tissue of breast-conserving surgery patients.

Abstract 2550: Tumor microenvironment (TME) is more suppressive in hepatocellular carcinoma (HCC) than in colorectal cancer liver metastasis (CRLM): CXCR4 antagonism as strategy to revert T regulatory cells (Tregs) suppressive activity

Abstract Background: Hepatocellular carcinoma (HCC) and colorectal cancer liver metastasis (CRLM) represent the two most occurring liver cancer. Aim of the project was to analyze the impact of TME on HCC versus CRLM. In particular, Tregs were evaluated in tumor (TT), peritumoral (PT) tissue and peripheral blood (PB) of liver tumor to identify immune mechanisms and therapeutic targets. Methods: Tregs were characterized on PB/PT/TT cell suspension from 40 HCC and 34 CRLM by flow cytometry (FOXP3, CD25, CD4, CD45RA, CTLA-4, CXCR4, PD-1, ICOS and ENTPD1). PB-, PT- and TT-Tregs were isolated by Dynabeads Regulatory T-cell kit and tested for the capability to inhibit CFSE-labeled autologous T effector (Teff) proliferation. Tregs were also treated with Pep R29, a new CXCR4 antagonists, before coculturing with Teffs. Quantitative real-time PCR was performed to assess the expression of cyto-chemokines in PT and TT samples (IL-15, Arginase, CCL5, CXCL12). Results: In HCC/CRLM-PB higher number of CXCR4 (p<0,001/p<0,05), CTLA-4 (p<0,001/p<0,05), PD-1 (p<0,001/p<0,05), ICOS (p<0,05/0,06% vs 0,03%) and ENTPD-1 (p<0,001/p<0,05) expressing Tregs was detected as compared to healthy donors (HD). In HCC/CRLM-TT higher number of FOXP3hiCD45RA-Activated Tregs subpopulation (p<0,01/p<0,05) and higher CXCR4-positive Tregs (p<0,05) were observed as compared to PT. Moreover, Tregs from HCC showed higher expression of ICOS (p<0,05) and ENTPD1 (p<0,05) as compared to CRLM. In terms of function, TT-Tregs exert a stronger immunosuppressive effect on Teffs proliferation than PT-Tregs (p<0,05) as shown by IL-35 cytokine secretion (150 vs 15 pg/mL/p<0,05). Interestingly, HCC-Tregs more powerfully inhibit Teffs proliferation at the tumor site and also in periphery. Expression of TME cyto-chemokines responsible of immunity promotion (IL-15) or inhibition (CXCL12, CCL5, Arginase) and tumor features, such as epithelial to mesenchymal transition (EMT) are under evaluation. Preliminary data showed higher CCL5, Tregs recruiting, in HCC as compared to CRLM suggesting a more suppressive TME in HCC. Based on high % of CXCR4-positive Tregs in HCC and CRLM (PB-HCC/CRLM: p<0,001 and TT-HCC/CRLM: p<0,01), ex vivo PB-Tregs were treated with Pep R29. CXCR4 targeting re-induced Teffs proliferation (PB-HCC -/+ Pep R29:12% vs 32%, p<0,001; PB-CRLM-/+ Pep R29:18,3% vs 49%, p<0,01) and decreased Tregs-derived IL-35 secretion (PB-HCC -/+ Pep R29: 106vs 16,5pg/mL, p<0,05; -/+ Pep R29:70 vs 0,6pg/mL, p<0,01). Conclusions: These data suggest a more suppressive TME in HCC compared to CRLM and highlighted CXCR4 antagonism as a potential strategy to overcome Tregs-mediated TME immunosuppressive function in HCC and CRLM patients. Citation Format: Daniela Castaldo, Sara Santagata, Giuseppina Rea, Maria Napolitano, Crescenzo D'Alterio, Rita Guarino, Vittorio Albino, Andrea Belli, Carmen Cutolo, Francesco Izzo, Stefania Scala. Tumor microenvironment (TME) is more suppressive in hepatocellular carcinoma (HCC) than in colorectal cancer liver metastasis (CRLM): CXCR4 antagonism as strategy to revert T regulatory cells (Tregs) suppressive activity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2550.

Abstract 5112: Investigating the use of circulating tumor DNA for sarcoma management

Abstract Circulating tumor DNA (ctDNA) has the potential to detect sarcoma recurrence and metastasis but requires highly sensitive methods to detect and quantify genetic variants present in very low quantities. Plasma was isolated from 20mL peripheral blood samples collected from over 400 pre-operative sarcoma patients, and matched tumor samples from surgical resection were frozen and stored. Cell-free DNA (cfDNA) extracted from plasma was quantified using qPCR, and the quality was assessed using capillary electrophoresis. A subset of these cases were selected for whole exome sequencing (WES). WES of bulk tumor and whole blood samples identified tumor-specific genetic alterations, which then serve as personalized biomarkers of tumor DNA in patient plasma. We previously showed that droplet digital PCR (ddPCR) can detect and quantify ctDNA, by targeting patient-specific variants. However, ddPCR is limited in that it can only investigate one tumor variant sequence at a time. The purpose of the present study is to investigate methods of targeting multiple tumor variants simultaneously, increasing the chances of detecting ctDNA in patient blood. To this end, four cases were selected for multiplex PCR (mPCR) followed by targeted amplicon sequencing. For each case, six to eight of the tumor variants identified by WES were selected as targets, and primers were designed to amplify these sequences concurrently by mPCR. The amplicons will then be sequenced to detect the tumor variants. Additionally, two of the four cases have plasma collected at two different time points. To assess the viability of this method as a way to monitor disease surveillance, these cfDNA samples will be compared to determine how the abundance and nature of ctDNA changes over time. To date, cfDNA has been extracted from over 100 cases, the majority of which were positive for cfDNA. For each of the cases whole exome sequenced, a variety of tumor-specific variations were identified. The variants chosen as targets were selected based on having the highest variant allele frequency (VAF), with priority being given to mutations that alter the protein coding sequence. Thus far, mPCR primers have been designed and optimized for four separate cases. Across all cases analyzed by amplicon sequencing, the variant sequences could be detected in the amplicons generated by mPCR of tumor DNA. Furthermore, amplicon sequencing was able to recapitulate the variant allele frequency observed in WES. This indicates that the mPCR successfully amplified the sequences of interest in the tumor DNA, and that the sequencing results are accurate. Furthermore, no tumor variants were detected in the amplicons generated from blood DNA, which is to be expected. The cfDNA amplicons for these cases will be sequenced in this manner to investigate the presence of ctDNA. If successful, the ability to detect ctDNA in plasma will be an important first step in developing a testing protocol for clinical use. Citation Format: Paige Darville-O'Quinn, Nalan Gokgoz, Kim M. Tsoi, Jay S. Wunder, Irene L. Andrulis. Investigating the use of circulating tumor DNA for sarcoma management [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5112.

Abstract 5758: Characteristic profile of blood-based ctDNA suggests angiogenesis and immune escape in hepatocellular carcinoma

Abstract Background: Circulating tumor DNA (ctDNA) is a type of DNA released from tumor cells captured in blood circulation system. It provides a noninvasive approach to interrogate a patient’s genomic landscape and actionable mutations with well application prospect. However, the difference of genomic profiles between tissue and ctDNA still brings confusions to related researches. Therefore, we tried to elucidate whether blood-based ctDNA has its own characteristic profile to reflect tumor status in hepatocellular carcinoma (HCC). Methods: Tumor tissue and blood samples from 118 HCC patients of Xiangya Hospital Central South University, were analyzed using NGS (panel on 147 gene). NGS data from a Chinese population extended cohort (385 tissues and 79 blood samples) of HCC were used to validate the mutation characteristics. Data from HCC cohorts of The Cancer Genome Atlas (TCGA) was used to analyze the disease-free survival (DFS) of different mutations. Results: In the hospital cohort, there were 9 high-frequency mutant genes in tissue and blood, including TP53 (74.58% vs 74.58%), MSH2(23.73% vs 8.47%), LRP1B (20.34% vs 23.73%), ATM (18.64% vs 13.56%), CTNNB1 (11.86% vs 13.56%) et.al which consistent with previous reports. There were 32 genes (74.04%) with a mutation frequency of more than 6% that were unique to blood samples. In extended cohort, the mutation profile was similar to the hospital cohort, in which 35 genes (52.24%) specific in ctDNA profile and 18 genes of them identical to the hospital cohort. Interestingly, when these ctDNA-specific genes were analysed by gene ontology, most genes were involved in angiogenesis. The prognostic analysis in ctDNA-specific genes showed that patients with higher level of MAP3K1 had worse DFS (p=0.028). In addition, the gene co-expression network analysis showed MAP3K1 and NOTCH1/2/3 expression had a significant positive correlation, that Notch family members were also belonged to the ctDNA-specific genes and associated with tumor immune escape. Conclusions: The results showed the mutation characteristic of tissues and ctDNA in HCC, and suggest that angiogenic and tumor immune escape related ctDNA-specific genes might be able to predict recurrence risk independently of baseline tissue samples. Citation Format: Lin Chia Yan, Liu Miao, Hu Xue, Huang Meng Li. Characteristic profile of blood-based ctDNA suggests angiogenesis and immune escape in hepatocellular carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5758.

Abstract 1849: Lipid metabolites as potential diagnostic biomarkers for stearoyl-CoA desaturase 1 targeting therapy in hepatocellular carcinoma (HCC)

Abstract New effective therapies should be developed for the treatment of hepatocellular carcinoma (HCC). One of the very promising strategies is blocking the activity of stearoyl-CoA desaturase 1 (SCD1). SCD1 is the enzyme responsible for de novo fatty acid (FAs) synthesis by converting saturated fatty acids (SFAs) into monounsaturated fatty acids (MUFAs). In our laboratory, we developed a novel, highly specific SCD1inhibitor - SSI-4. We have tested the biological activity of SSI-4 against different HCC cell lines and patient-derived xenografts (PDX) mouse models. Of the twelve tested HCC cell lines, four were highly sensitive to SSI-4 (IC50 1-50 nM). Other cell lines showed moderate or no sensitivity to SSI-4. We conducted analysis of the enzymatic protein levels involved in de novo fatty acid synthesis. Protein expression levels for these enzymes don’t predict response to therapy. To determine whether lipid metabolites predict response to SSI-4 therapy, lipidomic analysis was investigated. Our studies showed thatSSI-4 sensitive cells when exposed to SSI-4 had a higher baseline level of key lipid metabolites such as phospholipids (PL), diglycerides (DAG), diglycerides (TAG), ceramides (CE), and free fatty acids (FFA),compared to insensitive cells. We are currently validating these results in our in vivo models at the tumor tissue and blood levels. Additionally, mechanistic studies are in progress to determine if SREBP1c, an AKT regulated transcription factor known to regulate each of the fatty acid synthetic enzymes plays a role in HCC fatty acid synthesis and sensitivity to SCD1 inhibition. Citation Format: Justyna Joanna Gleba. Lipid metabolites as potential diagnostic biomarkers for stearoyl-CoA desaturase 1 targeting therapy in hepatocellular carcinoma (HCC) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1849.

Abstract 1681: Single cell RNA sequencing of primary Ewing sarcoma tumors and identification of circulating tumor cells in patient-matched peripheral blood samples

Abstract Ewing sarcoma (ES) is the second most common bone cancer in children, accounting for 2% of pediatric cancer diagnoses. Patients who present with metastatic disease at the time of diagnosis have a dismal prognosis, compared to the >70% 5-year survival of those with localized disease. Here, we utilized single-cell RNA sequencing (scRNA-seq) to characterize the transcriptional landscape of primary ES tumors, and to identify circulating tumor cells (CTCs) in peripheral blood at the time of diagnosis in order to further understand ES transcriptional heterogeneity and factors that drive metastasis. Methods: Viably frozen primary tumor and peripheral blood samples were obtained from 7 ES patients at the time of diagnosis and prior to the initiation of treatment. Tumors were dissociated into a single cell suspension and sorted for viability using FACS (fluorescence activated cell sorting) whereas peripheral blood samples were subjected to a size-based selection with the CellSieve microfiltration system. ScRNA-seq was performed using the 10xChromium platform and count matrices were generated using 10x Genomics Cell Ranger software. Quality control, integration, and cluster analysis was performed with Seurat. Results: Cluster analysis of integrated, primary tumor samples demonstrated that candidate ES cell clusters express FLI1 and ES marker NKX2-2 and cluster separately from immune cell populations. Additionally, using inferCNV, ES clusters were demonstrated to harbor chromosomal copy number alterations known to be associated with ES and that were previously identified clinically on preliminary pathology reports for each patient; further GSEA analysis showed significant overlap between published ES gene sets and genes upregulated in ES clusters. Further analysis of ES cells demonstrated that cell-cycle phase determined a cluster enriched in pro-proliferation gene signatures, and overall heterogeneity of expression of previously known therapeutic targets. Candidate ES CTCs were identified among the peripheral blood samples among clusters which corresponded with distinct immune cell populations. The candidate ES CTCs expressed NKX2-2 and demonstrated enrichment in oncogenic gene signatures. Conclusion: ScRNA-seq of primary ES tumors is feasible and demonstrates that ES tumor cells are largely homogeneous in nature; and candidate ES CTCs can be identified in peripheral blood at the time of diagnosis in ES patient and warrant further investigation as to their utility as a biomarker of metastatic disease. Citation Format: Sarah K. Nelson-Taylor, Avery Bodlack, Andrew Goodspeed, Amy Treece, Nathan Donaldson, Carrye Cost, Tim Garrington, Brian Greffe, Sandra Luna-Fineman, Jenna Sopfe, Masanori Hayashi. Single cell RNA sequencing of primary Ewing sarcoma tumors and identification of circulating tumor cells in patient-matched peripheral blood samples [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1681.

Abstract 45: Screening somatic genomic alteration for rare adult cancers: the SPECTA-ARCAGEN project of the EORTC/EURACAN

Abstract Introduction: Rare cancers are defined as having an incidence lower than 6 per 100,000 new cases per year. Despite advances in treatments and personalised medicine for cancer, rare cancers are often left behind due to limited knowledge regarding molecular characterization, heterogeneity of the population and poor access to screening platform or interventional trials.To bridge this gap, EORTC, in collaboration with EURACAN, has developed a project within the EORTC-SPECTA platform to assess the molecular landscape of rare cancers in Europe and the actionability of the different findings. Material and Methods: The pan-European Arcagen project is prospectively analyzing tumor material or blood of patient with rare cancers, using Foundation Medicine NGS panels, while collecting baseline and follow-up clinical data. Results: At the cut-off date of 10/10/2021, 629 patients were evaluable (baseline clinical data and successful molecular profiling performed by a Foundation Medicine test), molecular alterations were found in 620 patients, theoretically actionable ones in 421 patients. Importantly, approved treatment in the right tumor type was proposed in 58 patients (9.2%). Longer follow-up is needed to assess treatment adaptation and response. Most analyses were performed based on FFPE material (451 out of 629), but in case of low sample quality, low content of tumor cells, or old sample, rescue analysis was performed on liquid biopsy (177 out of 629). The main reason for switching to liquid biopsy was due to low FFPE tissue quality (90 out of 178). This rescue pathway was used on average in 28.1% of cases, ranging from 14.7% in salivary gland cancers to 48% for Cancer of Unknown Primary. Comparing per tumor type the molecular profile obtained either with FFPE or blood analysis, we noticed that SNVs are nicely detected in both compartments. However, copy number loss and amplifications are mainly detected when FFPE material is used. This is especially true for copy number loss of CDKN2A/B (cholangiocarcinoma, mesothelioma and salivary gland), or MTAP (mesothelioma), and MYC amplification (brain and rare skin cancer).As expected according to other datasets, some clonal hematopoiesis of indeterminate potential (CHIP) was observed on liquid biopsy, with DNMT3A being in the top ten mutations detected in the blood of patients with salivary gland tumors, endocrine tumors, Merkel cell carcinoma and uveal melanoma. Conclusion: This project highlights the clear need for more collaboration, research and clinical trials on rare cancers, to improve the treatment accessibility for targeted therapy for these patients. The use of liquid biopsy, despite its limitations, is an ideal candidate to gain understanding of a patient tumor molecular profile, while being non-invasive and easily performed longitudinally. Citation Format: Marie Morfouace, Aleksandra Stevovic, Julio Oliveira, Nicolas Penel, Julien Péron, Paolo G. Casali, Michael J. SECKL, Jourik A. GIETEMA, Wouter de HERDER, Lucjan WYRWICZ, Christelle de la FOUCHARDIERE, Lisa LICITRA, Nicolas GIRARD, Sophie PIPERNO-NEUMANN, Ellen KAPITEIJN, Ahmed IDBAIH, Jean-Yves Blay. Screening somatic genomic alteration for rare adult cancers: the SPECTA-ARCAGEN project of the EORTC/EURACAN [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 45.

Abstract 1258: Elevated serum DC-HIL is associated with poor response to immune checkpoint inhibitors (ICI) in metastatic renal cell carcinoma (mRCC)

Abstract ICI therapy has shown unprecedented clinical outcomes in mRCC patients, but durable responses have been limited by innate (never respond initially) and acquired resistance (initially respond but eventually develop cancer progression). Establishment of predictive biomarkers for response to ICI maximizes the therapeutic benefit. Since no such marker exists, we investigated DC-HIL/GPNMB as a possibility because we detected DC-HIL to be elevated in blood of patients with metastasis, as both a receptor on myeloid-derived suppressor cells (MDSC) and as a soluble form (sDC-HIL); and found DC-HIL to be relevant to ICI resistance in mice. We studied 27 mRCC patients treated with combination ICI (Ab to PD1/PDL1/CTLA4) and assessed tumor response (by RECIST) at 12 wk after therapy. Blood was collected before and after treatment and assayed by FACS for expression of DC-HIL and PDL1 on CD14+HLA-DRno/lo monocytic (mMDSC) and CD15+CD11b+ granulocytic MDSC (gMDSC). T-cell suppressor function was assayed by cell culture. We showed both MDSC subsets to express DC-HIL predominantly over PDL1. We also showed mMDSC to be more potent T-cell suppressors than gMDSC (85% vs. 58% suppression, p=0.012) and to also be more dependent on DC-HIL for such suppression (56% vs. 15% restoration of T-cell function after adding DC-HIL mAb, p=0.013). However, there was no significant difference in % DC-HIL+MDSC/PBMC between ICI responders and non-responders (p=0.15). We next examined the relationship between tumor response (at 12 wk) and blood sDC-HIL (by ELISA) in 31 mRCC patients. At baseline, non-responders (n=10) had a significantly higher sDC-HIL than responders (n=21) (14 ± 2.4 vs. 10 ± 4.6 ng/ml, p=0.004), but no difference in blood PDL1 or VEGF. Among initial responders (n=22), 50% maintained a positive response while the other half acquired resistance after 20 wk. Among initial non-responders (n=9), 77% never responded while 22% responded positively after 20 wk. We then analyzed changes over time in sDC-HIL levels after treatment (up to 60 wks) in individuals. Patients who responded positively to ICI at treatment inception and through 40 wk maintained blood sDC-HIL at levels lower than the median for all positive responders. Initial responders who acquired resistance (9 out of 11 patients) had low blood sDC-HIL that rose during cancer progression. All constant non-responders (n=7) had high blood sDC-HIL that remained high during the study. To our surprise, the 2 initial non-responders who responded positively after 20-40 wk of ICI displayed an abrupt rise in blood sDC-HIL during their positive response. Our results indicate blood sDC-HIL as a promising biomarker for prognosticating response to ICI in mRCC patients. Citation Format: Jin-Sung Chung, Vijay Ramani, Ponciano D. Cruz, Hans J. Hammers, Kiyoshi Ariizumi. Elevated serum DC-HIL is associated with poor response to immune checkpoint inhibitors (ICI) in metastatic renal cell carcinoma (mRCC) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1258.

Abstract 5400: Detection of satellite tumor cells in peritumoral edema in myxofibrosarcoma and undifferentiated pleomorphic sarcoma using targeted gene sequencing

Abstract Myxofibrosarcoma (MFS) and undifferentiated pleomorphic sarcomas (UPS) are soft tissue sarcomas with high local recurrence rates following resection. Common with these cancers is peritumoral edema, termed tumor tails, which our group has found could contain satellite tumor cells. A precise and accurate method of determining the presence of satellite tumor cells in this region is required to improve surgical planning, as the standard morphology-based technique is prone to misclassification. An emerging technology is targeted gene sequencing (t-NGS), a combination of whole genome or exome sequencing (WGS/WXS) comparing primary tumor and patient-matched blood, and digital droplet PCR (ddPCR). The objective of this study was to determine the viability of t-NGS in the detection of satellite tumor cells in the areas of edema surrounding MFS and UPS. A cohort of patients diagnosed with extremity MFS and UPS at Mount Sinai Hospital (Toronto, Canada) who underwent resection were retrospectively studied. Cases with available WGS/WXS data were used (n=8) to identify case-specific tumor-specific variants (TSV) to detect with the QX200 ddPCR platform. (n=4-5/case). Cases with a major TSV-positive droplet population in the tumor tails tissue indicated the presence of satellite tumor cells. Our quality analysis showed DNA above the ddPCR threshold of detection was extracted from the tumor tails of 6 of 8 cases. The ddPCR assay showed that a TSV-positive droplet population was detected within the tumor tails in 5 of 6 cases, while 1 of 6 cases showed no such population in the tumor tails. Overall, our results indicates that t-NGS has potential for TSV detection within tumor tails. A prospective study is underway to further test its validity and compare it with standard histology. The project aims to improve surgical planning by ensuring removal of satellite tumor cells while lowering the risk of wound complications. Targeted gene-sequencing results of all cases within this study Case # Histology Pre-op Radiation DNA Extraction Successful TSV Probes (n) TSV+ Tumor TSV+ Tails TSV+ Normal 1 MFS N Y 5 4 4 0 2 MFS N Y 3 2 2 0 3 MFS Y Y* (normal insufficient) 3 3 0 N/A 4 UPS Y Y 3 2 0 1 5 MFS N Y 4 4 1 0 6 UPS N Y 3 3 3 1 7 MFS Y N N/A N/A N/A N/A 8 MFS Y N N/A N/A N/A N/A Citation Format: Miguel Alfonso V. Principe, Nalan Gokgoz, Patrick Prochazka, Peter C. Ferguson, Simin Dewji, Jay S. Wunder, Irene A. Andrulis, Brendan C. Dickson, Kim M. Tsoi. Detection of satellite tumor cells in peritumoral edema in myxofibrosarcoma and undifferentiated pleomorphic sarcoma using targeted gene sequencing [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5400.

Association of DNA damage repair (DDR) mutations (mts) and clinical outcomes in CALGB 90601 (Alliance).

4521 Background: Platinum-based chemotherapy is the standard 1st-line therapy for metastatic urothelial cancer (mUC). C90601 was a randomized phase III trial testing gemcitabine and cisplatin (GC) with bevacizumab (B) or placebo (P) in patients (pts) with untreated mUC. Median overall survival (OS) for GCB vs GCP was 14.5 months (mo) vs 14.3 mo (p=0.14) and median progression-free survival (PFS) was 8 vs 6.7 mo, respectively. DDR mts have been implicated in response and survival in mUC and were investigated in this negative trial. Methods: C90601 enrolled 506 pts randomized 1:1 to GCB or GCP from 7/15/09-12/2/14, with stratification for prior chemotherapy and visceral metastases. Consenting pts submitted archival FFPE tumor specimens and blood for matched germline (g)DNA. Tumor and gDNA were sequenced by MSK-IMPACT, a 468-gene exon capture assay, to detect mts in select DDR genes. The proportional hazards model was used to correlate mts in the DNA helicase ERCC2 (pre-specified hypothesis) and additional DDR gene panels being explored in prospective trials in muscle-invasive disease with OS and PFS, adjusting for tumor mt burden and stratification factors. Mts were categorized as deleterious (del) or non-del using pre-defined published criteria. Results: 208 pts underwent DNA sequencing. Clinical features and PFS/OS were comparable to the 506-pt cohort. Median sequencing coverage was 497X. Median mutation count was 13.2 and 8.8 for DDR mt and wild-type tumors, respectively. A non-significant improvement in OS and PFS was seen in pts with ERCC2 mts (HR 0.70), but the 5.3% frequency of ERCC2 mts was lower than in historical series. Neither del mts (table) nor any mts in DDR genes were associated with PFS/OS. Conclusions: DDR mts were not associated with improved outcomes in C90601. The reliance on archival specimens, lower-than-expected ERCC2 mt frequency, small sample sizes, and tumor genomic heterogeneity may have influenced the predictive capacity of DDR mts in this cohort. Similar analyses are underway in pts who received neoadjuvant chemotherapy prior to cystectomy from completed prospective trials. Support: U10CA180821, U10CA180882, Genentech.[Table: see text]

Efficacy and safety of ASK120067 (limertinib) in patients with locally advanced or metastatic EGFR T790M-mutated non–small cell lung cancer: A multicenter, single-arm, phase IIb study.

9106 Background: ASK120067 (Limertinib) is a newly developed third-generation epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI) targeting both sensitizing EGFR and EGFR T790M mutations. This study aimed to evaluate the efficacy and safety of ASK120067 in patients with locally advanced or metastatic EGFR T790M mutated non-small cell lung cancer (NSCLC). Methods: This study was a single-arm, open-label, phase 2b study conducted at 62 hospitals across China. Patients with locally advanced or metastatic NSCLC with centrally confirmed EGFR T790M mutations in tumor tissue or blood plasma who progressed after first or second generation EGFR TKIs or with primary EGFR T790M mutations were enrolled. Patients received ASK120067 160mg orally twice daily, until disease progression, or unacceptable toxicity. The primary endpoint was objective response rate (ORR) assessed by Independent Review Committee (IRC) per RECIST1.1. Secondary endpoints included disease control rate (DCR), progression-free survival (PFS), duration of response (DOR), overall survival (OS), and safety. Safety was assessed according to CTCAE 4.03. Results: Between June 24th, 2019 and Feb 25th, 2021 301 patients were enrolled and received ASK120067 treatment. All patients entered the full analysis set (FAS) and safety set (SS). A total of 99 (32.9%) patients had central nervous system (CNS) metastases at baseline. By the data cutoff date on Sep 9th, 2021, 76 (25.2%) remained on treatment. The median follow-up time was10.4 (range 0.3-26.3) months. Based on FAS, the IRC-assessed ORR was 68.8% (95%CI 63.2%-74.0%) and DCR was 92.4% (95%CI 88.8%-95.1%). The median PFS was 11.0 (95%CI 9.7-12.4) months, median DOR was 11.1 (95%CI 9.6-13.8) months, and median OS was not reached (NR) (95%CI 19.7 months-NR). Objective responses were achieved across all pre-specified subgroups. For 99 patients with CNS metastases, the ORR was 64.6% (95%CI 54.4%-74.0%), median PFS was 9.7 (95%CI 5.9-11.6) months, and median DOR was 9.6 (95%CI 8.1-15.2) months. For 41 patients who had evaluable CNS lesion, the confirmed CNS-ORR was 56.1% (95%CI, 39.7%-71.5%) and median CNS-PFS was 10.6 (95%CI 5.6-NE) months. In SS, 289 (96.0%) patients experienced at least one adverse drug reaction (ADR), with the most common being diarrhea (81.7%), anemia (32.6%), rash (29.9%) and appetite decrease (28.2%). Grade ≥3 ADRs occurred in 104 (34.6%) patients, and the most common included diarrhea (13.0%), hypokalemia (4.3%), anemia (4.0%) and rash (3.3%). ADRs leading to dose interruption and dose discontinuation occurred 24.6% and 2% of patients, respectively. No ADR leading to death occurred. Conclusions: ASK120067 demonstrated promising efficacy and an acceptable safety profile for the treatment of patients with locally advanced or metastatic EGFR T790M mutated NSCLC. Clinical trial information: NCT03502850.

A first-in-human, phase 1b study to evaluate the safety, tolerability, pharmacokinetics, and anti-tumor activity of neoadjuvant use of ph-762 administered intratumorally in subjects with advanced melanoma.

TPS9608 Background: Antibodies targeting immune checkpoints such as PD-1 and CTLA-4 has shown significant benefit in late-stage melanoma. But further improvements in therapeutic options are still required. Two approaches for improving the outcome of immunotherapy with checkpoint inhibitors are neoadjuvant treatment and local intratumoral (IT) injection. IT immunotherapy uses the tumor as its own vaccine to activate the immune system, priming an anti-tumor immune response and generating systemic tumor responses, whilst minimizing systemic exposure and off-target toxicities. Neoadjuvant therapy provides the opportunity for preoperative disease shrinkage with the potential to improve surgical morbidity. There is currently no neoadjuvant standard of care for resectable, advanced melanoma patients. PH-762 is a potent RNAi molecule targeting PD-1 with structural and chemical modifications conferring properties suitable for IT administration, including an optimized cell and tissue uptake profile. Pharmacology studies show potent in vitro silencing of PD-1 associated with T cell activation, and robust, dose-dependent in vivo inhibition of tumor growth in syngeneic tumor models. Methods: The purpose of this study is to evaluate the safety of neoadjuvant use of PH-762 administered by IT injection in subjects with resectable stage IIIB/IIIC/IIID or IV melanoma, to determine the recommended Phase 2 dose, PK after IT injection, and potential immunologic and pathologic tumor responses. Study treatment constitutes of once weekly injections with PH-762 into one designated tumor lesion for 4 weeks prior to surgical excision at 5-6 weeks after the initial injection, with up to 5 dose levels tested in a serial fashion in cohorts of 3 or more subjects. Eligible subjects will have at least one resectable melanoma deposit that is large enough to allow IT injection, and that can undergo repeated biopsy. Subjects with active brain metastases, leptomeningeal disease, uveal melanoma, and auto-immune disease are excluded. The dose of PH-762 will be normalized to tumor volume to ensure an equivalent local dose (tumor tissue concentration). Post tumor excision, subjects will be followed-up for 6 weeks. Primary endpoint is to determine a safe dose of PH-762 assessed by incidence of Dose Limiting Toxicities (DLT) prior to tumor resection. Bayesian optimal interval (BOIN) design will be employed to evaluate escalating doses of PH-762 to determine the Maximum Tolerated Dose based on occurrence of DLT. Tumor changes will be evaluated per RECIST criteria (version 1.1 and iRECIST version adapted for use with IT therapy) and pathological response. Immunological response in tumor tissue and blood samples will be assessed as secondary endpoints. Enrollment commenced in February 2022. Clinical trial information: 2021-002859-10.

ACCRU-GI-2008: A phase II randomized study of atezolizumab (Atezo) plus a multi-kinase inhibitor (MKI) versus MKI alone in patients with unresectable advanced hepatocellular carcinoma (aHCC) who previously received atezolizumab plus bevacizumab (Bev).

TPS4170 Background: IMbrave150 is the first study demonstrating the benefit of anti-PDL1 in the frontline treatment of aHCC, and established Atezo/Bev as a new 1st line standard for aHCC. There is currently limited evidence to guide subsequent therapy for aHCC patients progressing on Atezo/Bev. ACCRU-GI-2008 is designed to determine the benefit of continuing Atezo into 2nd line and the safety of Atezo plus a MKI in patients with aHCC who previously received Atezo/Bev. The study is being conducted across 12 centers in the United States (ClincalTrials.gov#: NCT05168163). Methods: This study utilizes a 2:1 randomized phase II design where eligible patients will receive either Atezo/MKI (experimental arm) or MKI alone (control). Patients will be stratified according to the MKI choice (cabozantinib or lenvatinib, per physician’s decision), etiology of HCC (viral vs. non-viral) and alpha-fetoprotein level ( < 400 vs. > = 400 ng/mL). The major eligibility criteria are histological/cytological diagnosis or clinical diagnosis of HCC per the AASLD or WASL 2018 guidelines, has advanced disease not amendable to curative treatment, previously received and progressed on Atezo/Bev, has received only 1 previous line of systemic therapy (2nd line only), ECOG PS 0-1, Child Pugh Class A, adequate organ reserves and RECIST v1.1 measurable disease; previous MKI for advanced disease is excluded. The primary endpoints are overall survival (OS) and progression free survival (PFS). A total sample size of 122, with 89 PFS events, we will have 80% power to detect an improvement in median PFS from 4 to 7 months, assuming a one-sided significance level of 0.05. With approximately 84 deaths, we will have 80% power to detect an improvement in median OS from 10 to 18 months, assuming a one-sided significance level of 0.05. The overall one-sided significance level, for the study, is 0.1. An OS interim analysis will be conducted at 89 PFS events. Secondary endpoints include objective response, duration of response, and adverse events. Archival tumor and serial blood samples will be collected to evaluate for potential prediction biomarkers and mechanisms of sensitivity/resistance. Baseline and on-treatment tumor biopsy specimens will also be collected from the initial 10 patients of each arm. The study is approved by the ethics committee and enrollment to the study will be underway by Q2/3 2022. Clinical trial information: NCT05168163.

A phase I/II trial evaluating the safety and efficacy of eribulin in combination with copanlisib in patients with metastatic triple-negative breast cancer (TNBC).

TPS1128 Background: Metastatic (met) TNBC remains a clinical challenge with limited treatment options and inevitable chemoresistance. Aberrant PI3K pathway signaling is frequently observed in TNBC. Increasing evidence shows PI3K pathway activation maintains the stemness and chemoresistance of BC stem cells (CSCs), and PI3K inhibition sensitizes CSCs to chemotherapy (chemo). Eribulin (E), a non-taxane microtubule dynamics inhibitor, showed survival benefit in met HER2 negative BC. Preclinically, E impacts tumor vascular remodeling, inhibits epithelial-to-mesenchymal transition and metastasis – key mechanisms implicated in PI3K inhibition resistance. Copanlisib (C), a potent pan-class I PI3K inhibitor ( i), improved anti-tumor effect in E-sensitive and resistant TNBC patient-derived xenograft models, irrespective of PIK3CA/PTEN mutation (mut) status, when combined with E. This phase I/II study is aimed to determine the safety and efficacy of E+C in pts with met TNBC. Methods: This trial includes a phase I portion with the primary objective to determine the dose limiting toxicity (DLT) and recommended phase 2 dose (RP2D) of E+C, followed by a phase II randomized portion of E+C (at RP2D) versus ( vs) E with the primary objective of progression-free survival (PFS). Key secondary objectives include objective response rate (ORR) and clinical benefit rate (CBR) [phase I]; and ORR and CBR, by arm and by PIK3CA/PTEN mut status and assessment of treatment induced target engagement [phase II]. Key exploratory objectives include analysis of genomic, proteomic and metabolomic changes as potential response biomarkers in tumor tissue and blood. Key eligibility criteria include pts with: met TNBC who progressed on ≤5 chemo lines, including anthracycline/taxane (unless contraindicated), ECOG 0-1, adequate organ function and known archival tumor PIK3CA/PTEN mut status. Key exclusions: prior E or PI3K/mTOR/AKT i, grade ≥2 neuropathy, tumor AKT mut, congenital QT prolongation, and uncontrolled diabetes or hypertension. Phase I portion will follow a 3+3 design for E+C dose escalation to enroll 18 max pts, starting at E 1.1 mg/m2 IV and C 45 mg IV on days (D) 1/8 of 21-D cycle (C) (to E 1.4 mg/m2 and C 60 mg max). RP2D will be defined as the highest dose level at which at most 1 of 6 pts experience DLT during C1. 88 pts will be randomized (1:1) in the phase II portion to E+C vs E (1.4 mg/m2 D 1/8), stratified by PTEN/PIK3CA mut status. Response assessment by Response Evaluation Criteria in solid tumors (RECIST) v1.1 will occur every 9 weeks (+/-7 D). Tumor biopsy is required at baseline and C2D1-2, and optional at progression. A sample size of 88 achieves 80% power to detect PFS difference of median PFS 6.95 vs 4 months (corresponding to a hazard ratio of 0.5755) between the 2 arms, based on 1-sided two-sample log rank test at 0.1 α level. The phase I study is actively enrolling pts. Clinical trial information: NCT04345913.

FoRT 05-BEAT: A phase II randomized trial comparing atezolizumab versus atezolizumab + bevacizumab as first-line treatment in patients with PD-L1 high advanced/metastatic NSCLC.

TPS9145 Background: Several ongoing phase III studies are evaluating the efficacy of first-line atezolizumab in combination with chemotherapy in patients with non small-cell lung cancer (NSCLC). Among angiogenesis inhibitors, bevacizumab is approved as first-line therapy in combination with chemotherapy or in combination with erlotinib in patients with NSCLC harboring activating EGFR mutations. Recent evidence (Wallin 2016) suggests that the combination of atezolizumab and bevacizumab increases intra-tumoral CD8+T cells, suggesting that dual VEGF and PD-L1 inhibition improves antigen-specific T-cell migration. In addition, preliminary clinical data suggested a strong synergistic effects of bevacizumab with immune checkpoint inhibitors. There is therefore a strong rationale for investigating the combination of atezolizumab and bevacizumab in patients with advanced/metastatic NSCLC. The FoRT 05-BEAT is a multicenter, Italian, phase II, randomized study comparing atezolizumab monotherapy versus the combination of atezolizumab and bevacizumab in patients with chemo-naive metastatic NSCLC and high levels of PD-L1 expression. Methods: The trial was conducted in 35 Italian centers: chemotherapy naive metastatic NSCLC patients, with high levels of PD-L1 expression (PD-L1 TPS ≥50% or TC/IC 3 scoring) were randomly assigned to atezolizumab monotherapy (1200 mg every 3 weeks) or to the combination of atezolizumab (1200 mg every 3 weeks) and bevacizumab (15 mg/kg every 3 weeks).The primary endpoint is overall survival (OS) rate at 18 months. Secondary endpoints include response rate (RR), PFS, OS according to presence of bone and/or hepatic metastases. Safety considerations will be considered. Exploratory analysis of predictive biomarker on tumor tissue and blood samples has been planned. Sample size has been calculated assuming a 18 months OS of 50% in the atezolizumab arm. Therefore, a total of 186 patients is needed to detect an absolute improvement of 20%, thus obtaining a 18mOS of 70% in the combination arm, with a power of 80% at a significance level of 5%. Taking into account the percentage of patients lost-to-follow-up, the sample size has been increased by 10% (N = 206 patients, 103 per arm). At the drafting of this abstract, 57 patients have already been enrolled (47 randomized). Clinical trial information: NCT03896074.

Genomic landscape and homologous recombination repair (HRR) genes analysis in Chinese prostate cancer (PCa) patients.

e17020 Background: HRR gene mutations have been proven to be effective biomarkers for PARP inhibitor (PARPi) therapy in metastatic castration resistant PCa. The epidemiological data of genomic variation and the characteristic of HRR gene alterations in Chinese population is still insufficient. Methods: Tumor tissue samples (including surgical and biopsy samples) and blood samples from 250 pathologically confirmed PCa patients were collected and analyzed using next generation sequencing (733-gene panel). Tumor mutation burden (TMB) was defined as total number of somatic non-synonymous mutations in coding region. Homologous recombination deficiency (HRD) was evaluated by the sum of the loss of heterozygosity, telomeric allelic imbalance and large-scale state transition. Somatic or germline alterations (including copy number reduction, CNV-loss) of HRR pathway genes were classified as HRR-altered positive. Results: In total, the top three alteration frequency somatic genes were FOXA1 (24.9%), TP53 (15.2%) and MYC (13.6%) while the top three alteration frequency germline genes were DPYD (17.1%), UGT1A1 (13.2%) and SLX4 (4.3%). In HRR pathway genes, the top three alteration frequency somatic genes were CDK12 (9.7%), ATM (4.7%) and BRCA2 (3.9%) while the top three alteration frequency germline genes were BRCA2 (3.1%), PALB2 (1.9%) and BRCA1 (1.6%). 25.6% (64/250) cases carried HRR gene alterations. Among HRR-altered positive cases with HRD score, only 8.9% (4/45) carried more than one HRR gene alteration and most carried a single HRR gene alteration (40.0% [18/45] in CDK12, 17.8% [8/45] in ATM or BRCA2, 11.1% [5/45] in BRCA1 and 2.2% [1/45] in PALB2 or FANCA). 61 HRR gene alterations were detected in these 45 cases and the commonly alteration types of HRR genes were single base substitution (47.5%, 29/61). The alteration type of CNV-loss (14.8%, 9/61) was only detected in somatic samples. The HRD score of HRR-altered group was significantly higher than wild group (median HRD socre, altered-type vs. wild-type = 23.5 vs. 19.0, P= 0.0097). And there was no significant difference in HRD score between CNV-loss and other alteration types. Moveover, the TMB level of HRR-altered group tended to be higher than wild group (median TMB, altered-type vs. wild-type = 4.5 vs. 2.8 Muts/Mb, P= 0.5). Conclusions: The results illustrated the molecular characteristics of HRR gene in Chinese PCa and indicated that HRR gene alterations (including CNV-loss) were associated with higher HRD score. These data provide further evidence for the clinical management of PARPi personalized therapy.

Evaluation of tumor gene expression profile (GEP) in minority patients with stage IV non-small cell lung cancer (NSCLC).

e21043 Background: Differences in tumor gene expression profile (GEP) may contribute to disparities in lung cancer incidence and survival between racial and ethnic groups. For patients with stage IV NSCLC, tumor biomarker analysis is important for identification of potentially effective targeted therapies. However, despite advancements in biomarker testing and targeted therapy, minority patient enrollment in clinical trials remains low and it has been shown that minority patients are less likely to undergo next-generation sequencing (NGS) compared to Non-Hispanic Whites (NHW). As such, frequencies of tumor biomarkers in minority populations in the United States are not well-defined. This study aimed to determine the extent of tumor GEP diversity in minority patients vs NHW. Methods: This retrospective study included 196 adult patients with stage IV NSCLC that had a baseline NGS of tumor tissue or blood (liquid biopsy) performed between July 1, 2015 and June 30, 2021. Minority populations included Hispanic, African American, and Asian patients. Data collected utilizing the electronic medical record included patient demographics, detection of tumor biomarkers and treatment initiated. Descriptive statistics were calculated for all demographic and clinical variables. Comparisons of actionable epidermal growth factor receptor (EGFR) mutation frequency between NHW and minority patients were done using Chi-squared and Fisher’s exact tests. Results: Of the 196 patients evaluated, baseline characteristics included 46% male, 67% former smokers, 54% NHW, 31% Hispanic, 9% African American, 5% Asian, and 1% other. The median age at diagnosis was 66 ± 12 years. Actionable mutations (EGFR, KRAS, BRAF, ROS-1, ALK, MET, NTRK 1-3) were found in 50% of patients (see Table). The frequency of actionable EGFR mutations was higher in minority patients overall compared to NHW (39.3% vs 21.5%; p < 0.01). The frequency of actionable EGFR mutations was similar between NHW and Hispanic patients (21.5% vs 27.9%; p = 0.35), however differences were observed comparing NHW to African American (21.5% vs 50%; p = 0.02) and Asian (21.5% vs 90%; p < 0.01) patients, respectively. Conclusions: Among patients with stage IV NSCLC, the GEP of minority patients varied from that of NHW. Asian patients had the highest frequency of EGFR mutations, followed by African American and Hispanic patients, respectively. Larger studies are needed to validate these findings.[Table: see text]